This research project aims to define the organization of the adenovirus genome in terms of the location and function of the virus genes which control virus development during infection, and to define the requirements for adeno gene products in oncogenic transformation of rodent cells. Temperature-sensitive (ts) and host-range (hr) mutants of type 5 human adenovirus have been isolated, and these are being characterized genetically and physiologically. Many of these mutations have been genetically mapped, and have now been mapped physically by marker rescue with an improved transfection method using adeno DNA-protein complex. The hr mutations are defective for transformation and map within the left-hand early gene sequences. These have been partially characterized in terms of viral DNA synthesis and early viral mRNA synthesis; the results provide evidence that a very early viral gene product encoded by sequences between 1.5 to 3.5 units regulates expression of other early regions. The hr mutants are now being characterized in terms of early viral protein production, induction of cellular DNA synthesis and induction of transplantation immunity. The mutations of one set of early function, transformation-defective ts mutants (ts 36, ts 69, ts l49) have been mapped between 18.5 to 22.0 units on the genome. Additional early gene sequences have now been located in this region, and these ts mutants are being further characterized genetically and physiologically. The improved transfection method is being used to introduce specific fragments mutated or modified in vitro into the Ad5 genome.